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The blood-brain barrier (BBB) is a layer of cells that block most molecules from entering the brain in order to protect this sensitive organ from external invaders. It does its job so well that delivering treatments to the brain is extremely difficult and poses a great challenge to drug development.

Researchers from the University of Nebraska Medical Center, the University of North Carolina, and Lomonosov Moscow State University in Russia have found a potential solution. In early 2013, these collaborators announced that they were successful in delivering an enzyme to the site of cell death within the brain of a mouse experiencing poor motor function characteristic of Parkinson’s disease. The researchers accomplished targeted delivery through a method called Trojan therapy. This article will explain the concept of Trojan therapy, as well as its potential application for Huntington’s disease.

The method for drug delivery is named for the Trojan War tale, during which one army snuck past the defenses of their enemy by disguising themselves within large wooden horses. The horse was presented as a gift and when it was brought behind the city’s defenses the army used the element of surprise to defeat the enemy forces.

Trojan therapy works in a similar fashion. The blood-brain barrier serves as a fortress wall for the brain. In this case, researchers disguised potential treatments within macrophages, immune cells that are allowed entry past the blood-brain barrier, making it possible to deliver the targeted therapy straight to the cells that need it.

The scientists utilized an antioxidantenzyme (proteins that combat oxidative free radicals within the body) known as catalase, to prevent cell death within the mouse brain affected by Parkinson’s disease. They engineered macrophages to carry the genetic material for catalase past the blood-brain barrier and into the brain. Once inside the brain, the macrophages attached to dying neurons, stimulating signals that called for backup to repair these neurons.

In order for this type of therapy to function properly, blood-borne macrophages must be able to carry antioxidant proteins across the blood-brain barrier to affected areas of the brain. In this study, the scientists loaded the macrophages with nanozymes, a type of enzyme that was used to clear out free radicals from the brain.

Once the macrophage finds its way into the brain, there are three options for the nanozyme to reach its specific target in the brain:

3) Release of exosomes – large vesicles or storage pockets in the cell – containing the nanozyme. 1

Exosomes appear to be most promising for targeting cell death as research has shown they are excellent long-distance cellular transporters. They effectively transport many biological molecules involved in making proteins from the information contained in genetic material, including proteins, mRNA, and microRNA (Zomer et al, 2010). Researchers engineered macrophages to release exosomes containing DNA, mRNA, transcription factors (proteins that attach to DNA and controltranscription of genetic information to messenger RNA) and an encoded catalase protein to assist in reducing inflammation of the brain. In Parkinson’s disease mouse models, the researchers discovered that sustained production of catalase resulted in inflammatory and neuroprotective outcomes, meaning cell death and brain atrophy was not as likely in those mouse models treated by the macrophage Trojan therapy.

The expression of the encoded catalase included within the macrophage increased over time, while DNA levels remained constant, which has implications for the therapeutic efficacy. An antioxidant with greater positive outcomes might be able to be transferred to needy neurons through this mechanism of delivery.

The control group for this experiment utilized mouse models without inflammation. The experimental group included mice exhibiting motor signs of PD. After injecting the PD mice with the macrophages, noticeable changes occurred. The fluorescent markers used to tag the progress of the materials packaged within the macrophage showed thorough coverage of the brain, especially to regions of inflammation where cell repair support was needed. These mice were able to perform just as well as the control mice on balance tests after receiving the Trojan therapy.

Oxidative stress can lead to cell damage. Reducing excess free radicals is an attractive method of reducing inflammation in PD brains. In the past, these methods have proven unsuccessful due a lack of mechanisms to deliver therapeutic drugs across the blood-brain barrier. The research conducted by this collaboration of scientists is the first to find a potential route for overcoming these obstacles. In fact, this method might be more effective than traditional viral vector gene therapy used for these purposes.

The research by the University of North Carolina and Lomonosov Moscow State University highlights various issues faced by clinical researchers. As outlined in this article, ability to cross the blood-brain barrier plays a major role in therapy design. Additionally, as the brain is a very sensitive organ, it is difficult to gauge the toxicity thresholds for various treatments and medications. Exosomes, unfortunately, are not a silver bullet either. The efficiency of loading drugs into exosomes is a challenge, as well as the difficulty in targeting the exosome to a particular cell type or organ.

Scientist Andrew Feigin of Hofstra North Shore-LIJ School of Medicine is currently leading a human-based clinical trial in which he is trying to use virus-delivered gene therapy for PD. This method has more sustained impacts than macrophages, but is much harder to get through the brain as the immune system fights these foreign viruses trying to pass the blood-brain barrier.

Researcher Elena Batrakova of University of North Carolina aims to deliver growth factors into the brain utilizing macrophages, in order to stimulate the growth of neurons. She believes that macrophages are the solution to reversing neuronal death in patients with PD. To-date this research has only been conducted in mouse models. In order to start human trials, scientists need to prove the safety of this therapy, as well as assure this therapy can be repeated successfully. This process could take several more years before human trials begin.

While this study focuses on Parkinson’s disease, overcoming the challenge of effectively delivering medicine across the blood-brain barrier could be useful in preventing damage or repairing brain cells in any of the neurodegenerative disorders, including Huntington’s disease. If HD scientists develop drugs or treatments that could slow down or reverse neuronal death in Huntington’s disease patients, macrophage-based delivery, as pioneered in this research, could be a method for getting that medication across the blood brain barrier and into regions of the brain where the medication is needed. If a method of drug delivery, such as Trojan therapy, could be proven safe, effective, and broadly applicable, it would allow scientists to concentrate on drug development.